title: Multiple residues contribute independently to differences in ligand recognition between vesicular monoamine transporters 1 and 2.
abstract: The two closely related vesicular monoamine transporters (VMATs) 1 and 2 differ substantially in ligand recognition. The neuronal VMAT2 exhibits a higher affinity for monoamine substrates and in particular for histamine as well as a greater sensitivity to the inhibitor tetrabenazine than the nonneuronal VMAT1. The analysis of chimeric transport proteins has previously shown that two major domains, one spanning transmembrane domains (TMDs) 5-8 (TMD5-8) and the other, TMDs 9-12 (TMD9-12), are required for the high affinity interactions characteristic of VMAT2. Using site-directed mutagenesis to replace residues in TMD5-8 of VMAT2 with the equivalent residues from VMAT1, we now show that the sensitivity of VMAT2 to tetrabenazine requires Ala-315, and this interaction occurs independently of the interaction with residues in TMD9-12. The ability to recognize histamine as a substrate depends on Pro-237, and the contribution of TMD9-12 to histamine recognition appears to involve a common mechanism. In contrast, the replacement of many residues in TMD5-8 of VMAT2 with equivalent residues from VMAT1 improves the recognition of both serotonin and tryptamine, and these mutations show a dominant effect on the recognition of both tryptamine and serotonin over mutations in TMD9-12. The results indicate that different ligands interact through distinct mechanisms with the VMATs and that the recognition of each ligand involves multiple, independent interactions with the transport protein.
authors.affiliation: Department of Microbiology and Immunology, UCLA School of Medicine, Los Angeles, California 90024, USA.
authors.forename: J P
authors.initials: JP
authors.lastname: Finn
authors.forename: R H
authors.initials: RH
authors.lastname: Edwards
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001692:Biological Transport / Q000502:physiology; D019556:COS Cells; D006632:Histamine; D007700:Kinetics; D008024:Ligands; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009479:Neuropeptides; D017433:Protein Structure, Secondary; D012701:Serotonin / Q000378:metabolism; D013747:Tetrabenazine / Q000494:pharmacology; D014162:Transfection / Q000235:genetics; D014363:Tryptamines; D050492:Vesicular Biogenic Amine Transport Proteins; D050493:Vesicular Monoamine Transport Proteins
chemical_list: D008024:Ligands; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D009479:Neuropeptides; D014363:Tryptamines; D050492:Vesicular Biogenic Amine Transport Proteins; D050493:Vesicular Monoamine Transport Proteins; D012701:Serotonin; C030820:tryptamine; D006632:Histamine; D013747:Tetrabenazine